A functional musculoskeletal system depends upon the transmission of force from muscle to bone, thus requiring the coordinated development of muscle, cartilage and tendon. Little however, is known about the cellular and molecular aspects of tendon formation. Progress unraveling the signals that direct mesenchymal cells to become tenocytes is of particular importance in elucidation of the early events that shape the organization and coordinated growth of the musculoskeletal system. Moreover, a tendon inducing activity may be of considerable clinical importance both for improvements in the treatment of tendon injuries and in bioengineering efforts to induce tendon formation from stem cells. The recent finding that Scleraxis (Scx) a bHLH transcription factor is a unique marker of tenocytes and tendon progenitors facilitated studies of tendon development and a number of subsequent studies have implicated FGF signaling in the induction of tendon progenitors. Dramatic results in our laboratory now demonstrate an even more profound role for TGF( signaling in tendon induction and differentiation. Embryos doubly mutant for the TGF(2 and TGF(3 genes develop limbs in which no tendons can be detected. Moreover, TGF( signaling acts as a potent inducer of Scx, in organ culture and in tissue culture experiments, significantly exceeding the extent and intensity of Scx induction by FGF signaling in similar experiments. We propose to explore TGF( role in tendon formation in three separate groups of experiments. The first aim will address the hypothesis that all tendon tissues depend on TGF( signaling. The phenotype in axial and cranial tendons will be evaluated in TGF(2 &3 mutants and in mutants for a TGF( receptor (TGF(RII), in which all TGF( signaling is eliminated. We will further study the onset of the tendon phenotype in these mutants to establish the stage in which TGF( signaling is essential for tendon formation. The second aim will be to address the hypothesis that TGF( signaling is essential for later aspects of tendon formation as well. Inducible elimination of TGF(RII will enable exploration of later roles for TGF( signaling and establish if tendons are generated from a committed pool of early progenitors or through continuous recruitment of tendon cells by TGF( signaling to support tendon growth. Finally, the mechanism of Scx induction by TGF( signaling will be explored in tissue culture and in vivo;the activities of TGF( and FGF signaling will be compared and the possibilities of synergism or interdependence between these two signaling cascades will be explored using a combination of specific antagonists and mutant tissues. In addition, TGF( will be applied in utero using a retroviral system to evaluate the capacity of the cells induced by TGF( signaling to go beyond expression of Scx and cause the formation of ectopic or expanded tendons.